台湾林檎叶提取物不同极性部位抑菌活性高通量筛选及其谱效相关性分析

High Throughput Screening of Bacteriostatic Activity in Different Polar Parts of Extracts of Leaves of Malus doumeri and Spectrum-Effect Relationship Analysis

关键词:  台湾林檎  高通量  金黄色葡萄球菌  抑菌  谱效分析  物质基础

Key words:  Malus doumeri(Bois) Chev.  high throughput  Staphylococcus aureus  antibacterial  spectrum-effect relationship  material basis

摘要:

目的 实现台湾林檎叶醇提物不同极性部位抑菌活性快速、高效筛选和评价,研究不同极性部位HPLC指纹图谱与抑菌作用谱效关系,以期明确台湾林檎叶抑菌物质基础。方法 采用HPLC建立台湾林檎叶醇提物不同极性部位指纹图谱,采用金黄色葡萄球菌刃天青高通量抑菌筛选模型,对台湾林檎叶醇提物不同极性部位样品进行抑菌活性评价,计算不同浓度各极性部位的抑菌率,筛选台湾林檎叶抑菌活性部位。运用最小二乘拟合法来建立各极性部位共有峰与抑菌活性间的谱效分析,寻找对抗金黄色葡萄球菌作用贡献较大的成分;采用UPLC-Q-TOF/MS²技术对高贡献度的共有峰进行识别分析。结果 在台湾林檎叶醇提物不同极性部位指纹图谱中共确定15个共有峰,不同极性部位抑菌活性为乙酸乙酯部位>正丁醇部位/二氯甲烷部位>石油醚部位>萃余液。共有峰8、峰9、峰10、峰15等显示有较高的抑菌活性贡献度,分别鉴定为槲皮素-7-O-葡萄糖苷、异槲皮苷、根皮苷和白杨素。结论 台湾林檎叶抑菌活性部位为乙酸乙酯部位,谱效分析提示槲皮素-7-O-葡萄糖苷、异槲皮苷、根皮苷和白杨素4个成分为抑菌活性成分,是否存在其他微量抑菌活性成分还有待进一步研究。

Abstract:

OBJECTIVE To realize rapid and efficient screening and evaluation of antibacterial activity of different polar parts of the ethanol extracts of leaves of Malus doumeri, and to study the relationship between HPLC fingerprint and their antimicrobial activity, in order to determine the antibacterial material basis of leaves of Malus doumeri. METHODS HPLC was used to establish the fingerprint of different polar parts of the ethanol extracts of leaves of Malus doumeri. The anti-Staphylococcus aureus activities of different polar parts of the ethanol extracts of leaves of Malus doumeri were evaluated by high-throughput screening method with resazurin assay. Calculated the bacteriostatic rate of different polar parts at different concentrations to screen the antibacterial active parts of the ethanol extracts of leaves of Malus doumeri. Spectrum-effect relationship between the common peaks of each polar part and the antibacterial activity was established by least squares regression, selected the largest contribution of common peaks. Finally, effective peaks were identified by UPLC-Q-TOF/MS². RESULTS A total of 15 common peaks were picked up in the fingerprint of different polar parts of the ethanol extracts of leaves of Malus doumeri. The results showed that the order of antimicrobial activity was ethyl acetate fraction>n-butanol fraction/dichloromethane fraction>petroleum ether fraction>water fraction. Peak 8, peak 9, peak 10 and peak 15 showed high contribution to antibacterial activity, which were identified as quercetin 7-O-β-D-glucopyranoside, isoquercetin, phlorizin and chrysin. CONCLUSION The ethyl acetate fraction is the antibacterial active polar parts of the ethanol extracts of leaves of Malus doumeri. The HPLC spectrum-effect relationship suggests that quercetin 7-O-β-D-glucopyranoside, isoquercetin, phlorizin and chrysin are the antibacterial active components, but it remains to be further explored whether there are other trace antibacterial active ingredients in leaves of Malus doumeri.